title: Targeting p35/Cdk5 signalling via CIP-peptide promotes angiogenesis in hypoxia.
abstract: Cyclin-dependent kinase-5 (Cdk5) is over-expressed in both neurons and microvessels in hypoxic regions of stroke tissue and has a significant pathological role following hyper-phosphorylation leading to calpain-induced cell death. Here, we have identified a critical role of Cdk5 in cytoskeleton/focal dynamics, wherein its activator, p35, redistributes along actin microfilaments of spreading cells co-localising with p(Tyr15)Cdk5, talin/integrin beta-1 at the lamellipodia in polarising cells. Cdk5 inhibition (roscovitine) resulted in actin-cytoskeleton disorganisation, prevention of protein co-localization and inhibition of movement. Cells expressing Cdk5 (D144N) kinase mutant, were unable to spread, migrate and form tube-like structures or sprouts, while Cdk5 wild-type over-expression showed enhanced motility and angiogenesis in vitro, which was maintained during hypoxia. Gene microarray studies demonstrated myocyte enhancer factor (MEF2C) as a substrate for Cdk5-mediated angiogenesis in vitro. MEF2C showed nuclear co-immunoprecipitation with Cdk5 and almost complete inhibition of differentiation and sprout formation following siRNA knock-down. In hypoxia, insertion of Cdk5/p25-inhibitory peptide (CIP) vector preserved and enhanced in vitro angiogenesis. These results demonstrate the existence of critical and complementary signalling pathways through Cdk5 and p35, and through which coordination is a required factor for successful angiogenesis in sustained hypoxic condition.
authors.affiliation: School of Healthcare Science, Manchester Metropolitan University, Manchester, United Kingdom.
authors.forename: Alessandra
authors.initials: A
authors.lastname: Bosutti
authors.forename: Jie
authors.initials: J
authors.lastname: Qi
authors.forename: Roberta
authors.initials: R
authors.lastname: Pennucci
authors.forename: David
authors.initials: D
authors.lastname: Bolton
authors.forename: Sabine
authors.initials: S
authors.lastname: Matou
authors.forename: Kamela
authors.initials: K
authors.lastname: Ali
authors.forename: Li-Huei
authors.initials: LH
authors.lastname: Tsai
authors.forename: Jerzy
authors.initials: J
authors.lastname: Krupinski
authors.forename: Eugene B
authors.initials: EB
authors.lastname: Petcu
authors.forename: Joan
authors.initials: J
authors.lastname: Montaner
authors.forename: Raid
authors.initials: R
authors.lastname: Al Baradie
authors.forename: Francesca
authors.initials: F
authors.lastname: Caccuri
authors.forename: Arnaldo
authors.initials: A
authors.lastname: Caruso
authors.forename: Giulio
authors.initials: G
authors.lastname: Alessandri
authors.forename: Shant
authors.initials: S
authors.lastname: Kumar
authors.forename: Cristina
authors.initials: C
authors.lastname: Rodriguez
authors.forename: Jose
authors.initials: J
authors.lastname: Martinez-Gonzalez
authors.forename: Mark
authors.initials: M
authors.lastname: Slevin
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000704:Analysis of Variance; D015153:Blotting, Western; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects; D003124:Colorimetry; D051360:Cyclin-Dependent Kinase 5 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D003599:Cytoskeleton / Q000187:drug effects; D017931:DNA Primers / Q000235:genetics; D006801:Humans; D000860:Hypoxia / Q000188:drug therapy* / Q000209:etiology / Q000378:metabolism; D047468:Immunoprecipitation; D064326:MEF2 Transcription Factors / Q000378:metabolism; D046228:Microarray Analysis; D008856:Microscopy, Fluorescence; D020125:Mutation, Missense / Q000235:genetics; D018919:Neovascularization, Physiologic / Q000187:drug effects / Q000502:physiology*; D010455:Peptides / Q000378:metabolism / Q000494:pharmacology; D011554:Pseudopodia / Q000378:metabolism; D011687:Purines / Q000494:pharmacology; D034622:RNA Interference; D000077546:Roscovitine; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D020521:Stroke / Q000150:complications*
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C500695:CDCA5 protein, human; D018797:Cell Cycle Proteins; D017931:DNA Primers; D064326:MEF2 Transcription Factors; C578040:MEF2C protein, human; D010455:Peptides; D011687:Purines; D000077546:Roscovitine; D051360:Cyclin-Dependent Kinase 5; C495905:CDK5 protein, human